Recent studies of H-2L, a second D region gene in the murine major histocompatibility complex (H-2), have intensified our interest in pursuing research on the heterogeneity of H-2D region genes and gene products. These new findings include: 1) H-2L antigens have been found to display several unique properties which distinguish them from H-2K and D antigens; 2) molecular heterogeneity in addition to D and L has been reported in studies of certain D regions, whereas other D regions appear to determine a single gene product; 3) two putative H-2L genes have been independently cloned, each having a unique DNA sequence; and 4) a dominant idiotype has been found in the humoral response to Ld antigens. In the proposed study, we will extend our knowledge of H-2 antigens by exploiting knowledge already gained from the study of H-2L antigens. New alloantisera and monoclonal antibodies to D region-encoded antigens will be produced using mouse strain combinations such that only select determinants are recognized. To probe the B-cell recognition of D region encoded antigens, the idiotypes of these reagents will be compared using anti-idiotypic reagents produced to monoclonal antibodies. Precipitates formed with anti-H-2D reagents will be chemically compared by sequential precipitation, 2-D gels, peptide maps, and amino acid sequence analyses. Furthermore, these reagents will be used to: 1) explore the linkage relationship of D region loci; 2) quantitate the cell surface expression of D region loci; 2) quantitate the cell surface expression of D region-associated gene products; and 3) probe the T-cell recognition of specific D region determinants in allogeneic and virus-specific cytotoxic responses. Recognition of H-2 antigens by T-cells will also be investigated using cloned T-cell lines and T-cell hybridomas which recognize, or are restricted by, D region antigens. These homogeneous cell lines in conjunction with the above described reagents will be used to probe the role of idiotype in the T cell recognition of H-2 antigens. The information acquired in this study will help elucidate the genetics, evolution and structure of major histocompatibility antigens as well as probe the immunologic mechanisms of MHC restriction and T-cell cytotoxicity.